The epidermal growth factor receptor (EGFR), a member of the erbB receptor family, is widely expressed in human tissues and regulates important cellular processes including proliferation, differentiation and development (S. Yano et al., 2003, Anticancer Res. 23, 3639). EGFR over-expression occurs in a range of solid tumors and is associated with disease progression, resistance to chemotherapy and radiation therapy, and poor prognosis (Arteaga, 2001, J. Clin. Oncol., 19, 32). Consequently, the EGFR and its downstream signaling effectors are major targets of new therapeutics such as monoclonal antibodies and tyrosine kinase inhibitors (Arteaga, 2003, Semin. Oncol., 30, 3). However, clinical responses to existing anti-EGFR agents in cancer are often limited and thus a major research focus is the development of novel approaches to block EGFR expression and signaling (Bianco et al., 2005, Cancer, 12, 159).
MicroRNAs (miRNAs) are short, endogenous, non-coding RNA molecules that bind via imperfect complementarity to 3′-untranslated regions (3′-UTRs) of target mRNAs, causing translational repression of the target gene or degradation of the target mRNA (Bartel, 2004, Cell. 116, 281; Mattick & Makunin, 2005, Hum. Mol. Genet. 14, 121; Humphreys et al., 2005, Proc. Natl. Acad. Sci. USA. 102, 16961). mRNAs are involved in a range of processes that include development and differentiation (Chen et al., 2006, Nat. Genet., 38, 228), proliferation and apoptosis (Cheng et al., 2005, Nucleic Acids Res. 33, 1290), and have been implicated in cancer (Zhang et al., 2007, Dev. Biol. 302, 1). Interestingly, more than half of miRNA genes are located at sites in the human genome that are frequently amplified, deleted or rearranged in cancer (Calin et al., 2004, Proc. Natl. Acad. Sci. U.S.A. 101, 2999), suggesting that some miRNAs may act as oncogenes (‘oncomirs’, Esquela-Kerscher & Slack, 2006, Nat. Rev. Cancer 6, 259) or tumour suppressors (reviewed in Zhang et al., 2007, Dev. Biol. 302, 1). For instance, reduced expression of the let-7 family of miRNAs is associated with increased Ras oncogene expression and reduced survival in patients with non-small cell lung cancer (NSCLC) (Johnson et al., 2005, Cell 120, 635; Takamizawa et al., 2004, Cancer Res., 64, 3753). In contrast, increased miR-21 expression in gliomas (Chan et al., 2005, Cancer Res. 65, 6029), breast, colon, lung, pancreas, prostate and stomach cancers (Volinia et al., 2006, Proc. Natl. Acad. Sci. USA., 103, 2257) is associated with resistance to apoptosis, reduced chemosensitivity and increased tumor growth (Chan et al., 2005, Cancer Res. 65, 6029; Si et al., 2006, Oncogene, 26, 2799).
Computational approaches have been developed to predict miRNA targets. These methods have utilized criteria such as complementarity between target mRNAs and a ‘seed’ region within the miRNA thought to be critical for binding specificity, and conservation of predicted miRNA-binding sites across 3′-UTRs from multiple species (reviewed in Rajewsky, 2006, Nat. Genet., 38, 8; Maziere & Enright, 2007, Drug Discov. Today, 12, 452). It has been suggested that miRNAs may have the capacity to regulate hundreds or even thousands of target mRNAs (Lewis et al., 2005, Cell, 120, 15) and that much of this regulation might occur at the level of mRNA decay (Krutzfeldt, et al., 2005, Nature, 438, 685). Furthermore, specific miRNAs have the potential to regulate expression of several members of a signaling pathway or cellular process (Stark et al., 2003, PLoS Biol., 1, 60). However, the imperfect complementarity of miRNA:target interactions means that the identification and functional validation of true miRNA targets remains a major challenge.